p53通路微小RNA在卵巢癌细胞和浆液性卵巢癌组织中的表达及意义

目的 探讨微小RNA (miRNA) miR-449a、miR-449b和miR-192家族是否与miR-34家族一样可作为p53通路成员,在卵巢癌中发挥作用.方法 以DNA损伤药物阿霉素处理p53野生型卵巢癌A2780细胞,采用Western blot法检测A2780细胞中p53蛋白的表达改变,采用实时定量PCR法检测激活p53后miR-449a/b、miR-34a、miR-34b、miR-34c、miR-192和miR-194的表达.在P53突变型卵巢癌SKOV3.ipl细胞中过表达miRNA,检测SKOV3.ipl细胞的细胞周期改变.采用实时定量PCR法检测miR-449a/b、miR-34a、miR-34b、miR-34c、miR-192和miR-194在正常输卵管以及不同分级分期的浆液性卵巢癌组织中的表达.结果 在阿霉素处理后24h,卵巢癌A2780细胞中p53蛋白的表达明显升高,miR-34b、miR-34c、miR-449a/b的表达约升高19 ～ 21倍,但miR-192和miR-194的表达未见明显变化.转染miR-449b和miR-34c后,SKOV3.ipl细胞出现G1期阻滞.miR-449a/b在浆液性卵巢癌组织中的表达改变与miR-34b和miR-34c基本平行,它们在高级别和晚期浆液性卵巢癌组织中的表达水平,显著低于正常输卵管组织以及低级别和早期浆液性卵巢癌组织;miR-192、miR-194和miR-34a在卵巢癌组织中的表达未发现明显的特征,在正常输卵管组织中的表达也明显低于miR-449a/b、miR-34b和miR-34c.结论 miR-449a/b与miR-34b、miR-34c作为抑癌基因,在p53通路中发挥协同作用,它们的功能缺失与浆液性卵巢癌的发生、发展密切相关.

Expression and significance of microRNAs in the p53 pathway in ovarian cancer cells and serous ovarian cancer tissues

Objective The aim of this study was to investigate whether miR-449a,miR-449b and miR-192 family microRNAs play the same roles in p53 pathway as miR-34 family in ovarian cancer.Methods Wild-type p53 ovarian carcinoma cell line A2780 cells were treated with genotoxic agent adriamycin.The reactivation of p53 was detected by Western blot.The expression of miR-449a/b,miR-34a,miR-34b,miR-34c,miR-192 and miR-194 were detected by real-time quantitative PCR.Mutant p53 ovarian cancer cell line SKOV3.ipl cells were transfected with pre-microRNAs and the cell-cycle changes were detected.The expression level of miR-449a/b,miR-34a,miR-34b,miR-34c,miR-192 and miR-194 in serous ovarian carcinomas of varying grade and stage were compared with real-time PCR.Results The expressions of miR-449a/b,miR-34b and miR-34c were 19-fold to 21-fold elevated after p53 activation by genotoxic agent. Ectopic expression of miR-449b,as well as miR-34c,resulted in cell-cycle arrest in SKOV3.ipl cells.The expression of miR-449a/b was parallel with that of miR-34b,miR-34c,and were significantly lower in late stage and high-grade serous carcinomas than in the normal fallopian tube,early stage and low-grade serous carcinomas.The expression of miR-192,miR-194 and miR-34a did not show evident features in serous ovarian carcinomas and were much lower than miR-449a/b,miR-34b and miR-34c in normal fallopian tube.Conclusions As tumor-suppressor microRNAs,miR-449a/b,miR-34b and miR-34c cooperate and play important roles in p53 pathway. Their inactivation may contribute to the carcinogenesis and progression of serous ovarian carcinomas.